Fig. 1.

ISG 15 inhibits HIV-1 replication. (A) HIV-1 (NL43) proviral DNA and plasmids expressing ISG15, UBE1L, or UBP43 (1 μg) were cotransfected to 293T cells. Supernatants were collected at indicated times after transfection, and the levels of released virus were analyzed by RT assay. (B) 293T cells were cotransfected with plasmids expressing NL43 (2 μg) and ISG15 (0 to 10 μg), and 48 h after transfection; supernatants were collected for RT assay. The purified virions (Supporting Text) were analyzed by immune blotting with p24 antibodies. The levels of transfected DNA were kept constant by the inclusion of plasmids DNA. (C) U1.1 cells were first treated with TPA for 24 h and then infected with 301 ISG15 vector or 301 control vector that were VSV pseudotyped. At different times after TPA treatment, supernatants were collected and the virus levels determined by the RT assay. (D) Cell lysates from TPA-treated U1.1 cells, infected with 301-ISG15 viral vector (described in C) or the empty 301 vector, were analyzed at different times after TPA treatment by Western blot with HIV and ISG15 antibodies. The levels of Gag proteins in TPA-treated U1.1 cells infected with an empty 301 vector at 48 and 72 h are shown.